某猪场猪瘟病例的综合诊断及病原基因分析

2017年3月山东省莱芜市某猪场保育猪群发生以高热(40.8~41.5℃)、精神高度沉郁、食欲不振、皮肤出血潮红及耳尖发绀为主要症状的急性热性传染病。为明确引起此次疫情的致病因素,对患病猪进行了病理剖检、组织病理学检查、分子生物学检测、病毒分离与鉴定,对分离病毒进行了E2基因的分子克隆测序及同源性比对,最终确定该病由猪瘟病毒感染引起,并分离到1株猪瘟病毒(SDLW2017)。该病毒株E2基因全长1119个碱基(nt),共编码373个氨基酸(aa);相似性分析表明,SDLW2017分离株与其它参考株的核苷酸相似性介于82.1%~96.3%,氨基酸相似性介于88.5%~97.6%,与1.1亚群HCLV株的核苷酸和氨基酸相似性最低,分别为82.1%和88.5%,而与2.1亚群参考株的核苷酸和氨基酸相似性较高,分别为91.8%~96.3%和95.2%~97.6%,其中与2.1b亚型参考株(GXWZ02)的相似性最高,分别为96.3%和97.6%。     

应用定量RT-PCR对猪瘟兔化弱毒疫苗的质量评价研究

为进一步验证猪瘟兔化弱毒(HCLV)疫苗荧光定量RT-PCR(qRT-PCR)与兔体定型热试验之间存在正相关性,本研究利用qRT-PCR方法对278批次HCLV疫苗进行检测,分别检测其疫苗含量及牛病毒性腹泻病毒(BVDV)污染情况。结果表明,278批次猪瘟疫苗中有66批(24%)疫苗含量达不到规程标准,有42批(15%)疫苗存在BVDV污染;从所检疫苗中抽取6份qRT-PCR检测HCLV含量较低的疫苗,采用兔体定型热试验进行验证,两种方法结果吻合,表明qRT-PCR方法可以作为评价猪瘟疫苗质量的一种备选方法。     

规模化猪场脐带血猪瘟病原检测及遗传变异分析

为了了解某规模化猪场母猪猪瘟病毒(CSFV)带毒及病原遗传变异情况,应用荧光定量PCR对117份分娩母猪的脐带血进行猪瘟病原检测,对阳性样品进行CSFV E2基因RT-PCR扩增,研究CSFV遗传变异特性。结果表明,该猪场猪瘟病原阳性率为7.69%(9/117),5株CSFVE2基因之间的核苷酸同源性为97.8%~100%,与参考株的同源性为80.0%~97.5%,与C株和HCLV株的同源性分别为95.6%~96.7%、96.4%~97.5%;5株CSFVE2基因推导的氨基酸同源性为97.8%~100%,与参考株之间同源性为83.5%~96.7%,与C株和HCLV株同源性均为95.5%~96.7%。遗传进化树分析表明,5株CSFV E2基因属于1.1亚群,与疫苗株HCLV亲缘性最近;氨基酸变异位点分析表明,5株CSFV E2基因在位于抗原决定区内氨基酸发生较小程度的变异。表明该猪场通过胎盘屏障感染仔猪的猪瘟病毒可能来源疫苗株,这种现象应该引起重视。     

猪瘟病毒广西流行毒株E0基因的克隆与分析

应用RT-PCR方法对猪瘟病毒E0基因进行扩增、克隆及测序,用DNA Star分析软件对6株毒株与猪瘟兔化弱毒苗(HCLV)、石门(Shimen)株、Paderborn株、GXWZ02株的相应片段进行比较分析,构建了CSFV遗传进化树。序列分析表明,广西流行株与HCLV株、Shimen株、Paderborn株、GXWZ02株之间核苷酸的同源性分别为80.8%~81.5%、82.8%~83.3%、93.1%~94.2%、93.7%~94.2%,推导的氨基酸同源性分别为88.3%~89.7%、89.7%~91.1%、96.2%~97.7%、97.7%~99.1%;6株广西CSFV流行毒株与国内外已发表的14株病毒相应序列进行比较,构建遗传进化树,结果表明所比较的20株毒株分为2个基因群,广西流行毒株均属于基因群Ⅱ。本研究成功地对广西流行猪瘟病毒株的E0基因进行了测序分析,表明近年来广西流行毒株未发生较大的变异,但病毒株有远离疫苗株发展的趋势。     

2011年我国部分地区猪瘟病毒分子流行病学研究

本研究对2011年我国部分地区养猪场采集的疑似猪瘟病料组织样本209份,分别用FQ-RT-PCR和RT-nPCR两种方法检测猪瘟病毒(CSFV)。结果FQ-RT-PCR检测出55份猪瘟阳性,RT-nPCR检测出53份阳性,2种方法符合率达96.3%。将53份阳性产物进行测序及同源性分析,显示这53份流行株均为2.1基因亚型。所分离的流行株与疫苗株(HCLV)和石门株(SM)的核苷酸同源性分别为79.4%~83.1%和79.4%~82.5%。研究结果表明,目前我国部分地区的养殖场仍有猪瘟病毒的存在,且以2.1亚型为主;流行株与疫苗株同源性仍高于77.4%,结合免疫攻毒试验,说明目前使用的猪瘟疫苗依然有效。     

猪瘟疫苗中BVDV污染一步法RT-PCR检测方法的建立与应用

为建立一种快速检测猪瘟(CSF)疫苗中牛病毒性腹泻病毒(BVDV)污染的方法,试验根据GenBank上登录的BVDV 5'端非编码区基因组序列设计1对引物,经一步法RT-PCR反应条件的优化,建立了检测猪瘟疫苗中BVDV污染的一步法RT-PCR。结果表明:该方法对猪瘟病毒、猪繁殖与呼吸障碍综合征病毒、猪圆环病毒2型、猪伪狂犬病毒、牛副流感病毒3型、牛传染性鼻气管炎病毒的扩增结果均为阴性,对BVDV基因组RNA的检测灵敏度达0.1 ng,与细胞培养法的符合率达100%。应用该方法对猪瘟疫苗、牛血清等138份样品进行BVDV污染的检测,阳性污染率达11.59%。该一步法RT-PCR具有良好的特异性、敏感性、准确性和适用性,可用于猪瘟疫苗生产中血清、细胞、毒种、半成品等的质量控制及猪瘟疫苗临床应用中的质量检验。     

牛病毒性腹泻病毒TC株的分离鉴定及E0基因的序列分析

从新疆塔城地区某牛场采集患病犊牛全血,将RT-PCR方法鉴定为牛病毒性腹泻病毒(BVDV)阳性的样品接种到MDBK细胞,传代至第4代,出现典型病变(CPE)。通过间接免疫荧光试验,证明该分离病毒为BVDV,命名为BVDV-TC。设计引物对E0基因进行扩增和序列分析。所得片段与GenBank上已发表的BVDV毒株核苷酸序列的同源性为69.3%~93.8%,氨基酸同源性为73.6%~97.8%,TC株的遗传距离与VEDEVAC株最接近。BVDV-TC是第一个在新疆地区报道的1b亚型BVDV毒株,为新疆地区BVDV感染的防控提供依据。     

福建省一株猪瘟病毒流行毒株全基因组序列测定与分析

根据GenBank公布的猪瘟病毒(CSFV)标准参考毒株全基因组序列,设计11对引物,应用RTPCR方法分别对11个片段进行福建流行毒株CN-FJLY的全基因组扩增,与参考毒株进行序列分析比较。结果表明,福建流行毒株CN-FJLY基因组全长为12 296bp,与参考毒株SXCDK、HEBZ、GXWZ02、HNLY-2011和Zj0801的全基因组核苷酸同源性为93.0%～97.0%,而与1.1亚群的中国强毒Shimen、疫苗株HCLV全基因组核苷酸同源性为84.4%～85.3%。基于全基因组及E2基因构建的遗传进化树分析结果表明福建流行毒株CN-FJLY属于2.1b亚群,与Shimen株、HCLV疫苗株亲缘关系较远。与HCLV相比较CN-FJLY基因组变异较大的是5′UTR、Ems、E2、p7、NS2、NS5A、NS5B和3′UTR。综上说明福建省猪瘟病毒流行毒株正在向远离疫苗株的方向变异。     

22株鸭甲型肝炎病毒的分离鉴定及其VP1基因的序列分析

为了解鸭甲型病毒性肝炎的病原特征,对2017—2019年从山东省采集的27份疑似鸭肝炎病毒感染的病料采用鸡胚进行病毒分离,并进行RT-PCR鉴定,随后对分离株的VP1基因进行序列测定以及遗传变异分析。结果显示:从27份疑似病料中共分离出了22株病毒,8株为鸭甲肝病毒1型(DHAV-1),14株为鸭甲肝病毒3型(DHAV-3);VP1基因序列比对显示:8株DHAV-1与12株参考株的核苷酸相似性为91.7%~98.6%,氨基酸相似性为93.7%~97.5%,分离株的核苷酸相似性为97.2%~100%;14株DHAV-3与12株参考株的核苷酸相似性为89.4%~98.9%,氨基酸相似性为92.1%~100%,分离株的核苷酸相似性为93.9%~100%,属于Ⅰ型(GⅠ型)。研究结果将为鸭甲型病毒性肝炎的防控提供了参考。     

牛病毒性腹泻病毒实时荧光定量RT-PCR检测方法的建立

为了建立一种快速、特异、灵敏的检测牛病毒性腹泻病毒(BVDV)Taq Man实时荧光定量RT-PCR的方法,根据NCBI GenBank上已公布的BVDV、猪瘟病毒(CSFV)核酸序列进行比对,利用Oligo 6.71软件设计一对引物及一条探针,建立了检测BVDV的Taq Man实时荧光定量RT-PCR方法。通过对该方法的特异性、重复性及其敏感性进行相关试验,结果表明该方法检测出BVDV标准毒株Oregon C_(24)V为阳性,猪圆环病毒2型、猪伪狂犬病病毒、猪传染性胃肠炎病毒、猪流行性腹泻病毒、猪繁殖与呼吸综合征病毒和CSFV的检测均为阴性。对BVDV标准毒株最低检测量达到10~(-2.5) TCID_(50)。该方法检测同一样品重复进行8次检测,结果均一致,表明方法的重复性较好。应用该方法对6批猪瘟疫苗专用血清进行BVDV检测,阳性率为16.7%;猪瘟弱毒苗中未检出BVDV。建立的BVDV Taq Man实时荧光定量RT-PCR方法为生产无BVDV污染的猪瘟苗提供了有力的保障。     

Feline leukemia virus and feline immunodeficiency virus.

Ophthalmic manifestations of FeLV or FIV infection can occur in all ocular tissues and may be manifestations of direct viral effects or secondary to viral-related malignant transformation. Additionally, the manifestations of common feline ophthalmic pathogens may be more severe and poorly responsive to therapy because of the immunosuppressive effects of FeLV or FIV infection. Prompt diagnosis of underlying viral infection in cats with ophthalmic disease is paramount for accurate diagnosis and prognosis and is required for appropriate therapeutic decision making.     

Response of cattle persistently infected with bovine virus diarrhoea virus to bovine leukosis virus

Six cattle persistently infected with bovine virus diarrhoea virus (BVDV) and seronegative, and two control, virus negative seropositive cattle were inoculated with lymphocytes infected with bovine leukosis virus (BLV). The two controls produced a normal immune response to BLV, developing antibodies at four and five weeks after inoculation. Two of the six cattle persistently infected with BVDV developed a strong antibody response by six weeks after inoculation with BLV. Four developed a depressed response to BLV, characterised in three by a 'hooking' reaction in the immunodiffusion test which persisted in successive bleedings but was interspersed occasionally by a weak positive reaction. In one of these animals, a series of 'hooking' reactions was followed by a number of negative results. The fourth animal remained serologically negative until 16 weeks after inoculation when a 'hooking' reaction was observed followed by a series of negative results. BLV was isolated from all the cattle persistently infected with BVDV at 42 or 58 weeks after inoculation regardless of whether the serum samples gave negative, 'hooking', weak positive or positive reactions in the immunodiffusion test. BLV was consistently isolated from the nasal secretions of a steer which was BVDV negative but seropositive. The possibility of decreased immune responsiveness to BLV in animals persistently infected with BVDV should be considered when formulating regulations governing the testing of animals for freedom from BLV.     

Evaluating the protective efficacy of a trivalent vaccine containing Akabane virus,Aino virus and Chuzan virus against Schmallenberg virus infection

Schmallenberg virus (SBV), an arthropod borne pathogen, spread rapidly throughout the majority of Europe since 2011. It can cause a febrile disease, milk drop, diarrhea, and fetal malformation in ruminants. SBV, a member of the Simbu serogroup within the genus Orthobunyavirus, is closely related to Akabane virus (AKAV) and Aino virus (AINOV) among others. In the present study, 4 Holstein-Friesian calves were immunized twice four weeks apart with a multivalent, inactivated vaccine against AKAV and AINOV. Another 4 calves were kept as unvaccinated controls. All animals were clinically, serologically and virologically examined before and after challenge infection with SBV. AKAV- and AINOV-specific neutralizing antibodies were detected one week before challenge infection, while SBV-specific antibodies were detectable only thereafter. SBV genome was detected in all vaccinated animals and 3 out of 4 controls in serum samples taken after challenge infection. In conclusion, the investigated vaccine was not able to prevent an SBV-infection. Thus, vaccines for other related Simbu serogroup viruses can not substitute SBV-specific vaccines as an instrument for disease control.     

Detection of bovine virus diarrhea virus in a live bovine herpes virus 1 marker vaccine

In February 1999, 12 Dutch herds were vaccinated with a live bovine herpesvirus 1 vaccine from which bovine virus diarrhea virus (BVDV) could be isolated. All vaccine batches that were on the Dutch market and that had not yet reached the expiry date were tested for BVDV. In total, seven of 82 batches tested were found positive. Batch numbers TX3607, VB3914, VB3915, VB4046, TW3391, and TV3294 were positive for BVDV type 1, and batch number WG4622 was positive for BVDV type 2. This latter batch induced clinical signs of BVDV in an animal experiment with susceptible animals.     

登革病毒及其所致疾病的研究概况

登革病毒所致疾病包括登革热、登革出血热及登革休克综合征，是全球分布最广、发病最多的一种虫媒传染病．近年发病呈上升趋势，严重威胁人类健康。文章概述了登革热病原学、流行病学、发病机制、临床表现、诊断进展、治疗、预防和控制等。     

Persistent bovine virus diarrhoea virus infection in a bull

Investigation of a sight defect in a pedigree bull, born as a result of artificial insemination and ovum transplantation, led to the finding that the animal was persistently infected with bovine virus diarrhoea virus. Virus was cultured from blood and from nasal and ocular swabs and was present in semen in high titre. At necropsy, virus was cultured from a wide range of tissues. The pathological findings are described and discussed as are the potential hazards of such infections.